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BP: Fachverband Biologische Physik

BP 15: Proteins II

BP 15.3: Vortrag

Mittwoch, 28. März 2012, 15:45–16:00, H 1058

Photo-cycle dynamics of photo-activated adenylate cyclase (nPAC) from the amoeboflagellate Naegleria gruberi NEG-M strain — •Alfons Penzkofer¹, Manuela Stierl², Peter Hegemann², and Suneel Kateriya³ — ¹Fakultät für Physik, Universität Regensburg, Universitätsstrasse 31, D-93053 Regensburg, Germany — ²Institut für Biologie/Experimentelle Biophysik, Humboldt Universität zu Berlin, Invalidenstrasse 42, D-10115 Berlin, Germany — ³Department of Biochemistry, University of Delhi South Campus, Benito Juarez Road, New Delhi 110021, India

nPAC comprises a BLUF domain (blue light sensor using flavin) and a cyclase homology domain (CHD). The nPAC gene was expressed heterologously in E.coli and the photo-dynamics of the nPAC protein was studied by optical absorption and fluorescence spectroscopy. Blue-light exposure of nPAC caused a typical BLUF-type photo-cycle behavior (spectral absorption red-shift, fluorescence quenching, absorption and fluorescence recovery in the dark). Additionally, time-delayed reversible photo-induced one-electron reduction of fully oxidized flavin (Fl_ox) to semi-reduced flavin (FlH^.) occurred. Furthermore, photo-excitation of FlH^. caused irreversible electron transfer to fully reduced anionic flavin (FlH⁻). A photo-induced electron transfer from Tyr to flavin (Tyr^.+ - Fl^.− radical ion-pair formation) caused H-bond restructuring responsible for BLUF-type photo-cycling and permanent protein re-conformation enabling photo-induced flavin reduction by proton transfer. Some photo-degradation of Fl_ox to lumichrome was observed. A model of the photo-cycle dynamics of nPAC was developed.